The studies from this laboratory have established the existence of a multigenic isozyme system for human phosphofructokinase (PFK). Three separate genetic loci are found to code for three distinct subunits, i.e., M (muscle), L (liver), and P (platelet). A tissue may express one or more genes; random polymerization of the resultant subunits result in the formation of one or many homo- or heterotetrameric isozyme(s). Our studies have also characterized the genetic defect of Tarui's disease (inherited PFK deficiency associated with hemolysis and myopathy) at the molecular level and established differential in-vivo stability of fetal red cell isozymes. The proposed research project aims at confirming and extending our preliminary observations regarding the following: (1) the molecular structures of three platelet and 10-12 fibroblast isozymes; (2) the existence of the as yet unidentified "T" or thyroid-type subunity (or T4) in several tissues; (3) the differential gene expression exhibited by lymphocytes on blast transformation in-vitro and in-vivo; (4) immunochemical analysis of isozymes using monospecific antibodies; (5) assignment of PFK genes to chromosomes and determination of linkage, if any; (6) elucidation of the biochemical mechanism(s) by which PFK deficiency leads to premature cell-death. These goals will be approached by the following means: (a) direct chromatographic resolution of various species from a given tissue; (b) confirmation of their subunit structures by molecular hybridization and immunoprecipitation techniques; (c) production of monoclonal antibodies by the use of secretory lymphocyte hydridomas; (d) use of somatic cell hybridization techniques to assign PFK genes to chromosomes; (e) studies of red cell glycolysis in steady state and changes induced therein by controlled conditions. These studies are expected to shed more light on the basic properties, genetic control and physiologic functions of this key regulatory enzyme of glycolysis. These will also aid our understanding of the physiological significance of PFK isozymes in control of carbohydrate metabolism at the cellular level in health and disease.